Methods for inhibiting the expression of a target gene in cells, tissues, or individuals include an approach in which double-stranded RNA is introduced into the cells, tissues, or individuals. By this introduction of double-stranded RNA, mRNA having homology to the sequence is degraded such that the expression of the target gene is inhibited. This effect is called “RNA interference” or “RNAi”. RNA interference was originally reported in C. elegans (see e.g., Non Patent Reference 1) and then also reported in plants (see e.g., Non Patent Reference 2).
Double-stranded RNA consisting of 21-nucleotide sense and antisense strands having a 2-nucleotide overhang at the 3′-end (small interfering RNA: siRNA) has been reported to have an RNA interference effect in cultured cells of vertebrates (see e.g., Non Patent Reference 3). siRNA is considered to be useful for the identification of gene functions, screening of cell lines suitable for useful substance production, regulation of genes involved in disease, etc., but it is characteristically degraded easily by RNase (see e.g., Non Patent Reference 4).
Since a double-stranded polynucleotide such as siRNA or modified siRNA is a molecule having a molecular weight on the order of 13,000, water solubility, and electric charge, a delivery technique such as a transfection reagent is generally used for allowing the double-stranded polynucleotide to permeate a cell membrane (see e.g., Non Patent Reference 5). Particularly, liposomes are widely used in the delivery of nucleic acid molecules by encapsulating a nucleic acid molecule such as plasmid DNA into a liposome to form a nucleic acid lipid particle (see e.g., Non Patent Reference 6). Also, a liposome containing a cationic lipid has been reported to be able to deliver siRNA into cells by forming a nucleic acid lipid particle through mixing with the siRNA (see e.g., Patent Reference 1). The cationic lipid, however, is a non-biological component. In this respect, a cationic lipid that can be used at a low concentration has been demanded. A dimethylaminovaleric acid derivative (Patent Reference 1), a dimethylaminobutyric acid derivative (Patent Reference 2), a dimethylaminoethylcarbonate derivative (Patent Reference 3), or the like is known as the cationic lipid.
The present inventors have conducted diligent studies to obtain a lipid particle consisting of a cationic lipid that can encapsulate therein a nucleic acid such as a double-stranded polynucleotide (e.g., siRNA), DNA, or an antisense oligonucleotide and can be used at a low concentration. As a result, the present inventors have completed the present invention by finding a novel cationic lipid and further finding a nucleic acid lipid particle comprising the novel cationic lipid that can encapsulate therein a nucleic acid molecule, can be used at a low concentration, and permits a high level of delivery into cells.